Predatory marine snails in the genus Conus have venoms that are rich in neuropharmacologically active peptides (Armishaw et al., 2005; Wang et al., 2004; Livett, et al., 2004; Lewis, 2004; Terlau et al., 2004). There are approximately 500 species in Conus, and among those that have been examined so far, a conserved feature is the presence of α-conotoxin peptides in their venom. α-Conotoxin peptides are highly disulfide cross-linked peptides with C1-C3 and C2-C4 disulfide bonds.
Due to high sequence variability of their non-cysteine residues, α-conotoxins are extremely diverse and each Conus species has a unique complement of α-conotoxin peptides. α-Conotoxin peptides are synthesized as large precursors, and the mature toxin is generated by a proteolytic cleavage toward the C-terminus of the precursor. In contrast to the variable inter-cysteine sequences of the mature toxins, the precursors and the genes encoding them are quite conserved both among α-conotoxin peptides in a given Conus species and from species to species.
α-Conotoxin peptides have generally been shown to be nicotinic acetylcholine receptor (nAChR) antagonists (Mcintosh, et al., 1999; Janes, 2005; Dutton et al., 2001; Arias et al., 2000). nAChRs are a group of acetylcholine gated ion channels that are part of the ligand gated ion channel superfamily (Karlin, 2002; Gotti et al., 2004). They are pentamers of transmembrane subunits surrounding a central ion conducting channel. Many different subunits have been identified, and most fall into two main subfamilies (the a subunits and the β subunits). The subunits can associate in various combinations in the receptor pentamers, leading to a diverse family of receptor subtypes. Most of the subtypes contain subunits from both the α and β subunit families, e.g., the human adult muscle subtype contains two a subunits and a β subunit (in addition to a δ and an ε subunit) and the α3β2 subtype is composed of α3 and β2 subunits. nAChRs that are composed of only a subunits are the α7 and α9 subtypes (homopentamers) and the α9α10 subtype (an all α heteropentamer). Phylogenetic analysis shows that the α7, α9, and α10 subunits are more closely related to each other than they are to other nAChR subunits (Le Novere, et al., 2002; Sgard, et al., 2002).
The α9 and α10 nAChR subunits are expressed in diverse tissues. In the inner ear α9α10 nAChRs mediate synaptic transmission between efferent olivocochlear fibers and cochlear hair cells (Sgard, et al., 2002; Elgoyhen, et al., 1994; Elgoyhen, et al., 2001). The α9 and α10 subunits are also found in dorsal root ganglion neurons (Harberger, et al., 2004; Lips, et al., 2002), lymphocytes (Peng, et al., 2004), skin keratinocytes (Arredondo, et al., 2002; Nguyen, et al., 2000; Kurzen, et al., 2004), and the pars tuberalis of the pituitary (Sgard, et al., 2002; Elgoyhen, et al., 1994; Elgoyhen, et al., 2001). In addition, the α9 nAChR subunit is active in breast cancer (Lee, et al., 2010a; Lee, et al., 2010b; Linnoila, 2010). α-Conotoxin peptide RgIA (RgIA; GCCSDPRCRYRCR; SEQ ID NO:1) has been shown to block α9α10 nAChR (Ellison, et al., 2006). Certain analogs of RgIA have also been shown to block α9α10 nAChR (US 2009/0203616, US 2012/0220539, and WO 2008/011006).